Determination of serum levels of unesterified lanosterol by isotope dilution-mass spectrometry.
The synthesis of 2H3-labelled lanosterol is described. This compound was used for assay of unesterified lanosterol in serum by isotope dilution-mass spectrometry. After addition of a fixed amount of internal standard (150 ng) to a fixed amount of serum (250 microliters) the steroids were extracted with chloroform and subjected to Lipidex 5000 chromatography. The fraction containing lanosterol was eluted with methanol. This fraction was converted into trimethylsilyl derivative and subjected to mass spectrometric analysis with selected monitoring of the ions at m/z 498 (molecular ion of unlabelled lanosterol) and m/z 501 (molecular ion of 2H3-labelled lanosterol). A standard curve was used for quantification of lanosterol in serum. Under the conditions employed the coefficient of variation was less than 4%. In a recovery experiment the maximal difference between expected and found value was about 5%. Sera from 10 healthy subjects were found to have a mean concentration of lanosterol of 225 ng/ml with a SD of 63 ng/ml. Using a less accurate method for analysis of lanosterol we have shown previously that there is a high correlation between the hepatic HMG CoA reductase and the absolute and relative concentration of lanosterol in serum (concentration of lanosterol relative to cholesterol). When using the same (old) sera as in the previous investigation this could not be confirmed, most probably because of the degradation of lanosterol as a consequence of prolonged storage and/or freezing and thawing. It appears that serum lanosterol must be analysed as soon as possible after the collection of the blood sample in order to obtain accurate results.(ABSTRACT TRUNCATED AT 250 WORDS)
Lund
E
E
Department of Clinical Chemistry, Karolinska Institutet, Huddinge Hospital, Sweden.
Olund
J
J
Björkhem
I
I
D000328:Adult; D015415:Biomarkers; D002784:Cholesterol / Q000096:biosynthesis; D002792:Cholestyramine Resin / Q000494:pharmacology; D003903:Deuterium; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry / Q000379:methods*; D006801:Humans; D006903:Hydroxymethylglutaryl CoA Reductases / Q000096:biosynthesis; D007810:Lanosterol / Q000097:blood*; D008297:Male; D008875:Middle Aged; D012016:Reference Values
D015415:Biomarkers; D002792:Cholestyramine Resin; D007810:Lanosterol; D002784:Cholesterol; D003903:Deuterium; D006903:Hydroxymethylglutaryl CoA Reductases